Lactobacillus sakei Probio65 extract (P65-CFS) was tested for its safety and efficacy in inhibiting SARS-CoV-2 replication. Viral load quantification by RT-PCR showed that the P65-CFS inhibited SARS-CoV-2 replication in human embryonic kidney (HEK) 293 cells in a dose-dependent manner, with 150 mg/mL being the most effective concentration (60.16% replication inhibition) (p < 0.05). No cytotoxicity was inflicted on the HEK 293 cells, human corneal epithelial (HCE) cells, or human cervical (HeLa) cells, as confirmed by the 3--2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. The P65-CFS (150 mg/mL) also reduced 83.40% of reactive oxidizing species (ROS) and extracellular signal-regulated kinases (ERK) phosphorylation in virus-infected cells, both of which function as important biomarkers for the pathogenesis of SARS-CoV-2. Furthermore, inflammatory markers, including interferon-α (IFN-α), IFN-ß, and interleukin-6 (IL-6), were all downregulated by P65-CFS in virus-infected cells as compared to the untreated control (p < 0.05). It was conclusively found that L. sakei Probio65 showed notable therapeutic efficacy in vitro by controlling not only viral multiplication but also pathogenicity; this finding suggests its potential to prevent severe COVID-19 and shorten the duration of infectiousness, thus proving useful as an adjuvant along with the currently available treatments.In the three years since the first outbreak of COVID-19 in 2019, the SARS-CoV-2 virus has continued to be prevalent in our community. It is believed that the virus will remain present, and be transmitted at a predictable rate, turning endemic. A major challenge that leads to this is the constant yet rapid mutation of the virus, which has rendered vaccination and current treatments less effective. In this study, the  Coronavirus Disease-2019 (COVID-19), rapidly recognized as a pandemic, triggered a global public health emergency with a death toll of approximately 15 million between 2020 and 2021. The predominant clinical manifestations exhibited by COVID-19 patients are related to respiratory and gastrointestinal tract disorders. The virus that causes the disease—SARS-CoV-2—is an enveloped, single-stranded RNA virus that transmits via aerosols (respiratory droplets suspended in the air) when prolonged face-to-face contact is maintained, or when a person is exposed to contaminated surfaces [Lactobacillus and Bifidobacterium, as well asdepleted gut diversity, and an abundance in populations of pathogens [Lactobacillus rhamnosus GG, live Bacillus subtilis, and Enterococcus faecalis) significantly reduced the need for ventilators [Previous studies have established that a dysbiotic intestinal microbiome can cause pulmonary dysfunction by modulating the host’s immune response through the gut–lung axis . In paraathogens . A studyathogens . In addiathogens . Therefotilators .Interestingly, a study by Bousquet et al. pointed out that the low COVID-19 death rates in Eastern Asia and Central Europe may be attributed to the high level of consumption of fermented food, particularly fermented vegetables, which is common to these regions . The stuL. plantarum Probio-88 (plantaricin E and F) in the inhibition of SARS-CoV-2 replication and in immune regulation, using an in vitro and in silico approach [Due to their numerous therapeutic implications for intestinal health and other areas related to the gut–organ axis, probiotics can dynamically regulate the hosts’ health ,9. They approach . In thisLactobacillus sakei Probio65, which is isolated from Kimchi, a type of fermented cabbage from Korea, has previously been reported to exert broad antimicrobial properties, as well as displaying antiviral activity against avian influenza virus (H9N2), immune modulation, and the ability to reduce inflammation [L. sakei Probio65, our research used an in vitro approach to determine whether the strain could affect the SARS-CoV-2 antiviral immune response.The strain ammation ,14. The L. sakei Probio65, a patented strain previously isolated from Kimchi, a traditional Korean fermented cabbage. The bacteria were cultured at 37 °C for 18 h in modified de Man–Rogosa–Sharpe (MRS), as previously reported [The bacterium used in this study was reported . The strg for 10 min. The supernatant was used as the virus stock, aliquoted, and stored at −80 °C. Virus stocks were quantified by titration.The SARS-CoV- 2 strain used in this study was obtained from an oropharyngeal swab specimen of a patient who had tested positive for coronavirus disease 2019 (COVID-19). All infection experiments were performed under biosafety level 3 (BSL-3) containment conditions with enhanced respiratory personal protection equipment. The virus was propagated in HEK 293 cells at 90% confluency for 48 h. Cell supernatant was collected after infection and clarified by centrifugation at 200× L. sakei Probio65 (P65-CFS) was produced by culturing the bacteria in modified MRS broth at 37 °C for 24 h, which was then centrifuged at 3500 rpm for 15 min. The clear supernatant was collected, neutralized to pH 7.0, and filter-sterilized using a syringe filter (pore size 0.22 µm). Finally, the supernatant was concentrated by freeze-drying and reconstituted with sterile distilled water to 10% of its original volume. The extract was stored at −20 °C for subsequent analysis.The CFS of 2. The cells were seeded in 96-well plates (4 × 104 cells per well) and incubated for 24 h. Then, the medium was replaced with DMEM containing different concentrations of P65-CFS  for 48 h at 37 °C and 5% CO2. Sterile PBS was used instead of P65-CFS in DMEM for the untreated control. At the end of the incubation period, the cells were washed in PBS, added to 50 μL of 5 mg/mL 3--2,5-diphenyl-2H-tetrazolium bromide (MTT) solution, and incubated for 45 min at 37 °C. All supernatant was aspirated; then, the purple formazan crystals were dissolved in isopropanol and the absorbance was measured at 570 nm. Percent viability was defined as the relative absorbance of P65-CFS-treated cells as compared to the control cells.The safety and cytotoxicity of P65-CFS to the human corneal epithelial (HCE), HEK 293, and Hela cell lines were tested using the MTT assay. In brief, each cell type was cultured in complete medium (Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) , penicillin (100 U/mL), and streptomycin (100 µg/mL), and incubated at 37 °C in a humidified atmosphere of 5% CO4 cells per well, and incubated for 24 h. The attached cells were treated with 100 μL of the culture medium for virus infection (DMEM supplemented with penicillin (100 U/mL) and streptomycin (100 μg/mL) + P65-CFS  + SARS-CoV-2 virus (0.1 multiplicity of infection (MOI)). MOI was calculated based on the viral titer of SARS-CoV-2, which was quantified by RT-PCR as RNA copies/mL of the virus in respect to the number of cells seeded per well. For the positive control, the same amount of the virus was first heat-inactivated at 95 °C for 3 min prior to its addition to the cells. The virus was allowed to infect the cells for 2 h at 37 °C, and then the cells were rinsed three times with PBS to wash off any unabsorbed virus. Thereafter, the cells were continuously maintained in 100 μL of the same culture medium containing the P65-CFS in the absence of virus for 48 h. Positive and negative control samples were inoculated in the same manner, but the P65-CFS was replaced with 0.1% DMSO (vehicle) and phosphate-buffered saline (PBS), respectively [Prior to infection, HEK 293 cells were seeded in 96-well plates at a density of 4 × 10Virus-induced changes in intracellular ROS levels were quantified using a DCFDA/H2DCFDA cellular oxidation kit, as previously described . HEK 293SARS-CoV-2-infected and P65-CFS-treated HEK293 cells were washed three times in PBS and fixed in 4% formaldehyde for 10 min. The cell membranes were permeabilized using 0.2% Triton X for 15 min, added to an image-iT FX signal enhancer, and then blocked with 5% goat serum diluted in 1X PBS. Cells were incubated at 4 °C with a primary antibody specific for p-ERK (1:100) for 12 h, after which the cells were incubated with secondary antibodies for another 1 h. Three PBS washes were performed between every step. The cell nuclei were counterstained with DAPI , washed with PBS, and finally photographed with a fluorescent microscope equipped with a digital camera .TM  was used to determine the RNA concentration and purity prior to the RT-PCR experiment. Briefly, 1 µg of RNA was added to a reaction mixture in a total volume of 25 µL containing random primers, and reverse-transcribed at 72 °C for 5 min and 37 °C for 60 min. The resulting cDNA was used in the real-time PCR experiment by adding 2 μL of the cDNA, 10 pmol of each gene-specific primer ), and the Power SYBR® Green PCR Master Mix  [−ΔΔCT method.Total RNA was extracted from SARS-CoV-2-infected and P65-CFS-treated HEK293 cells using Trizol reagent . NanoDropMA, USA) . The expWe investigated the cytotoxic effects of P65-CFS on the human embryonic kidney (HEK) 293 cells, human corneal epithelial (HCE) cells, and human cervical (HeLa) cells at varying concentrations using the MTT assay . IncreasOnce it had been proven safe, the inhibitive effect of P65-CFS on SARS-CoV-2 replication was then evaluated by culturing the virus on HEK 293 cells at 0.1 MOI. According to the treatment scheme described in Coronaviruses induce mitochondrial stress in the infected cells, which exacerbates the intracellular accumulation of reactive oxidizing species (ROS) and subsequently cell damage. In this study, when the HEK293 cells were infected with SARS-CoV-2, ROS were evidently accumulated in the intracellular compartment, as indicated by the green fluorogenic dye dichlorofluorescein (H2DCFDA) A. Using Extracellular signal-regulated kinases (ERK) are one of the three main groups of mitogen-activated protein kinases (MAPKs), which regulate cell cycles, gene transcription, and post-transcriptional regulation. During coronavirus infection, they play an important role in apoptosis and cell survivability ,18. The p < 0.05) (p < 0.05)  . The tre < 0.05) .While COVID-19 may now seem to have transitioned into a much more controllable phase, compared to the last three years, it is now a major global public health burden. This burden disproportionately affects the elderly population, who are at increased risk of severe disease, hospitalization, and death. We believe that there is still an urgent need for the development of new therapeutics that are safe, relatively cheap, and easily distributable to a wide range of populations. Our aim was to investigate the possibility of natural alternatives, such as probiotics, which have been known for decades to possess immunomodulating and antiviral properties. Probiotics have also been well reported as a therapeutic agent or adjuvant for vaccines in treating viral infections .L. sakei Probio65, at a concentration that was not cytotoxic to the HEK 293 cells, significantly reduced SARS-CoV-2 replication by up to about 60%. Our results showed that this reduction in virus replication is accompanied by a concomitant reduction in ROS production. ROS play an important role in regulating the expression of some transcription factors and signal transduction molecules in living organisms. While they are important for cell cycle replication, ROS are considered damaging if they are produced in excessive amounts. During SARS-CoV-2 infection, viral proteins induce endoplasmic reticulum (ER) stress and mitochondrial dysfunction, which eventually triggers increased levels of ROS through inflammasomes [In this study, we observed that treatment using the extract of mmasomes ,22.As ROS continue to accumulate, the redox balance becomes disorganized, activating a series of immune responses. In particular, ROS have been reported to activate the STAT/IL-6 axis, which is responsible for regulating the release of cytokines to the site that is inflamed or has been exposed to the virus . In mostL. sakei Probio65 successfully lessened ERK phosphorylation, suggesting that the extract has the ability to affect the virus–cell signaling process during infection.The increased levels of ROS during viral infection have also been reported to interfere with the ERK  pathway, which regulates vital cellular functions such as cell cycle progression, gene transcription, and post-transcriptional regulation . The ERKL. sakei Probio65 led to the reduced expression of these cytokines in comparison to the SAR-CoV-2-infected cells, suggesting that the extract prohibited viral replication in the cells, and could also regulate the homeostasis of immune activation during viral infection.During a viral infection, our innate immunity plays an essential role as the first line of defense in sensing and activating antiviral mechanisms. As soon as an infection takes hold, the pattern-recognition receptors (PRR) detect the viral components and stimulate cytokine production as part of the innate immune response of the cells ,30. AmonLactobacillus plantarum Probio-88. In the current study, meanwhile, the chemical composition of the extract from L. sakei Probio65 has not yet been elucidated. Nevertheless, L. sakei Probio65 is a safer and well-characterized probiotic strain.The SARS-CoV-2 strain used in this study was not sequenced. However, the focus of this study was on the replicative effect of the virus on the host’s cells, and we believe the objective of the study was achieved without the strain being sequenced. However, at present, new variants are emerging due to mutations on the spike proteins, which are more closely related to the virus’s attachment to and entry into the host cells, and were not the focus of our study. Additionally, there are no significant differences between the results of this study and those of our previous study , except L. sakei Probio65 for COVID-19 through its inhibition of viral multiplication and also of the subsequent pathogenicity. Our findings support its potential as a microbiome drug that might be used to prevent severe COVID-19 infection and to shorten the duration of infectiousness; it may therefore be useful as an adjuvant along with the currently available treatments.In light of the continued prevalence of COVID-19 and the emergence of new variants, such as the XBB subvariant of Omicron detected in Saudi Arabia in October 2022, it is essential to develop potent antiviral compounds for use against SARS-CoV-2. This study highlights the therapeutic efficacy of


Treatment of acromegaly resistant to first generation somatostatin analogues (first gen-SSA) is often difficult. We aimed to investigate the role of partial response and resistance to first gen-SSA in the choice of second line treatments and their outcomes.A retrospective and multicenter study was conducted on 100 SSA-resistant acromegaly patients and treated with Pasireotide Lar (Pasi-Lar), Peg-V in monotherapy (m-Peg-V) or in combination with first gen-SSA (c-Peg-V).p = 0.01) and with not-invasive adenomas (p = 0.009), c-Peg-V therapy in younger patients (p = 0.001), with invasive adenomas (p = 0.02), Pasi-Lar was in invasive adenomas (p = 0.01) and in patients partially responsive to first-gen SSA (p = 0.01). At the last follow-up, 68 patients (68%) reached the acromegaly control: 22 with m-Peg-V (32.4%), 23 with c-Peg-V (33.8%) and 23 with Pasi-Lar (33.8%). Patients non-responsive to c-Peg-V had higher IGF-I levels  and required higher Peg-V dosage  as compared to responsive patients . All patients responsive to Pasi-Lar were partially responsive to first gen-SSAs (p = 0.02).Thirty-three patients (33%) were treated with m-Peg-V, 36 (36%) with c-Peg-V and 31 with Pasi-Lar (31%). According to logistic regression, m-Peg-V was chosen in older patients (Our data showed that c-Peg-V and Pasi-Lar are chosen for the treatment of invasive tumors. The partial response to first gen-SSA seems to be the main determinant for the choice of Pasi-Lar and positively predicts the treatment outcome. Acromegaly is a chronic, systemic and complex disease, with an increased risk of mortality due to the disease per-se and to the increased prevalence of systemic complications . The reaThe surgical removal of the pituitary adenoma and the medical treatments with first generation somatostatin analogues (first gen-SSAs) represent the cornerstones in the management of acromegaly. According to the most recent meta-analysis, first gen-SSAs can induce the control of hormonal hypersecretion from 40% to 65% of acromegaly patients . As for An epidemiological, retrospective, and multicentre study was conducted on clinical data of acromegaly patients followed-up at four hospital-based endocrinology centres in Rome, Italy. Data were retrospectively collected. Data were anonymized, recorded on electronic forms by physicians involved in the patients’ care for each endocrine centre and sent to the Coordinating Centre for analysis. Data collection was conducted between November 2018 and November 2019. Patients involved in the study signed an informed consensus.Inclusion criteriapatients with active acromegaly at the moment of prescription of second line treatments, such as Pegvisomant or Pasireotide Lar;partial response or resistance to first generation SSAtreatment with PEG-V in monotherapy (m-Peg-V) or in combination with first gen-SSAs (c-Peg-V) or treatment with Pasireotide Lar for at least 12 consecutive months.Exclusion criteriaHistory of radiotherapy within 10 years before the prescription of Pasireotide LAR or Pegvisomant;treatment with Pegvisomant or Pasireotide Lar in clinical trialsduration of treatment with Pegvisomant or Pasireotide Lar in clinical trials shorter than 12 months.Partial response to first generation SSA was defined in patients who did not achieve the normalization of GH and IGF-I but with a significant decrease of GH and IGF-I levels (at least by 50% in comparison to pre-treatment levels) and/or in patients with a tumor shrinkage of at least 20%. Poor response or resistance to first generation SSA was defined in patients without a significant decrease of GH and IGF-I levels (less than 50% in comparison to pre-treatment levels) and in the absence of tumor shrinkage . ResponsBaseline was to the visit/day of the first dose of Pegvisomant or Pasireotide LAR. The treatment choice for Pegvisomant or Pasireotide LAR was based on the phsyician’s clinical judgment in the absence of reference guidelines, which were first published in October 2018 [GH secreting pituitary adenomas were considered as invasive in the cases of invasion of cavernous sinus and/or of the III ventricle. Cavernous sinus invasion was considered for tumors that extended laterally to the lateral tangent of the intra- and supra-cavernous internal carotid artery and in cases of total encasement of the intra-cavernous carotid artery (grade 3 and 4 of Knosp classification) . InvasioThe primary objective of the study was to investigate if the partial response and the resistance to first-generation SSA impacted on the choice of second line treatments in acromegaly patients. As secondary objectives, we investigated if other clinical determinants  may impact in choosing second line therapies and may predict the response to second line treatments.patients treated with Pegvisomant in monotherapy;patients treated with Pegvisomant in association to first generation SSA;patients treated with Pasireotide Lar.The primary endpoint of this study was the number of patients treated with Pegvisomant or Pasireotide LAR. According to the treatment choice, patients were divided in three groups:The secondary endpoint of this study was the number of patients who reached the control of acromegaly, in the three groups of treatments. At the last follow-up, patients were defined as controlled if the IGF-I values were in the reference ranges for age and gender (at least in two consecutive measures) and random GH was below 1.0 ng/mL . PatientThe patients’ cohort was described in its clinical and demographic features using descriptive statistics techniques. Normality of continuous variables was checked using the Kolmogorov-Smirnov test. Quantitative variables were expressed as median and range and qualitative variables as absolute and percentage frequency. Chi square test (or Fisher exact test when necessary) and Mann Whitney non-parametric tests were used to compare categorical and quantitative un-paired data. Bonferroni correction was applied as appropriate. The variables that reached a statistical significance at the univariate analysis entered the logistic regression. The analyses were performed using SPSS software version 24.0 for Windows.The study population included 100 patients. Clinical features of the study population are summarized in Table Thirty-three patients were treated with Pegvisomant in monotherapy (33%), 36 with Pegvisomant plus first gen-SSAs (36%) and 31 with Pasireotide Lar (31%). The initial median dose of Pegvisomant was 10 mg/daily (IQR: 5 mg/daily) in m-Peg-V group and 15 mg/daily (IQR: 10 mg/daily). Fifteen patients were treated with Pasireotide Lar 40 mg/monthly and the remaining 16 patients were treated with Pasireotide Lar 60 mg/monthly (respectively 48.4% and 51.4%).Thirty-six patients were affected by systemic arterial hypertension (36%), 29 by cardiomyopathy (29%), 28 patients by diabetes mellitus (28%), 17 patients by obstructive sleep apnoea syndrome (17%), 42 patients by nodular thyroid disease (42%), 19 patients by osteoporosis/osteomalacia (19%) and 13 patients (13%) had an history of second neoplasia . During the treatments, a transaminase elevation  was observed in 7 patients (7% of cases: 4 were on treatment with m-Peg-V and 3 on c-Peg-V), an injection-site reaction in 2 patients (2%) and a regrowth of residual tumour in 3 patients (2.8% of cases: one case was observed for each group of treatment).p = 0.002) and to patients treated with Pasireotide Lar (44 years IQR: 27 p = 0.04). Moreover, GH values at acromegaly diagnosis were higher (median 6.4 ng/mL IQR: 5.5) in patients treated with c-Peg-V as compared to those treated with m-Peg-V (median 4.3 ng/mL IQR: 3.3 p = 0.01) and to those treated with Pasireotide Lar (median 4.4 ng/mL IQR: 5.5 p = 0.005). In parallel, we found that GH values after treatment with first gen-SSA were higher in patients treated with c-Peg-V (median 4.8 ng/mL IQR: 14) as compared to those of patients treated with m-Peg-V .As showed in Table p = 0.007 and 71% p < 0.001) with respect to patients treated with m-Peg-V (21.2%). Moreover, we found that patients treated with c-Peg-V were more frequently affected by obstructive sleep apnoea syndrome (OSAS) (36.1%), with respect of patients treated with m-Peg-V  and Pasireotide Lar . Patients treated with Pasireotide Lar instead were more frequently affected by osteoporosis/osteomalacia (38.7%) and nodular thyroid disease (61.3% p = 0.02), with respect to those treated with m-Peg-V .Patients treated with c-Peg-V or Pasireotide Lar more frequently carried an invasive pituitary adenoma (respectively 52.8% p = 0.07 and 37.9% p = 0.01).In addition, as showed in the Fig. p: 0.02, r = 0.351), as shown in Fig. At the last follow-up visit, 68 patients reached the acromegaly control (68%), without any significant difference among groups of treatment. We analysed the clinical, hormonal and morphological features to identify potential predictors of second line medical treatments, as showed in Table p = 0.437). Instead, the expression of the SSTR5 diffuse and membranous (grade 2 and 3 of Volante score) in all 12 cases , as shown in Fig. According to the multicenter and retrospective design of our study, data on the SSTR expression were available only for 27 cases (9 treated with c-Peg-V and 18 treated with Pasireotide Lar). In the group of the 12 patients responsive to Pasireotide Lar, the expression of the SSTR2A was focal and cytoplasmatic (grade 0 and 1 of Volante score) in 6 cases (50%) and diffuse and membranous (grade 2 and 3 of Volante score) in the remaining 6 cases .Among the 31 patients treated with Pasireotide Lar, all patients with controlled diseases were considered partially responsive to first-generation SSA, as shown in Fig. p < 0.001), as showed in Supplementary Fig. 2.To better explore the efficacy of Pasireotide Lar in short-term follow-up, we also investigated the percentage of patients who reached the acromegaly control at 6 months of treatment. To reach this secondary objective, we analysed the efficacy of Pasireotide Lar at 6 months of treatment in the present cohort of 31 patients treated with Pasireotide Lar and in a small group of other 10 acromegaly patients treated for at least 6 months, but that did not reach the 12 consecutive months of treatment. As for consequence, a group of 41 acromegaly patients was finally retrospectively analysed. Thirty-four were partially resistant to previous therapy with first-gen SSAs and the remaining 7 patients were completely resistant to first-gen SSA. At six months of treatments with Pasireotide Lar, 29 patients (70.7% of cases) reached the control of acromegaly. In this sub-analysis, we confirmed that Pasireotide Lar was preferred in patients that were partial resistant to first generation SSA . Moreover, we found that the complete resistance to SSA remained a risk factor for a poor response to Pasireotide Lar  were treated with m-Pegvisomant in monotherapy, 36 (36%) with Pegvisomant plus first gen-SSAs and 31 patients with Pasireotide Lar (31%).This study showed for the first time that the partial response achieved by the treatment with first gen-SSA may oriented physicians into choosing Pasireotide Lar as a second line treatment. In fact, we found that 94% of patients treated with Pasireotide Lar were considered partially responsive to first gen-SSA. Recently, Muhammad et al. . Data onAs described in a previous multicenter cohort of acromegaly patients resistant to first gen-SSA , the actp = 0.004) and with Pasireotide Lar . Also, these results supported and underlined the theory that patients treated with c-Peg-V were affected by a more aggressive/severe disease. This result confirmed our previous experience that an IGF-I superior to 3 x ULN may negatively predict the treatment outcome of Peg-V therapy [In this study, we reported active acromegaly in 32 patients at the last follow-up: 11 patients were on treatment with m-Peg, 13 with c-Peg-V and 8 with Pasireotide Lar. We found that patients with active acromegaly during c-Peg-V therapy carried significantly higher IGF-I x ULN levels at the start of second line therapies , with respect to those treated with m-Peg-V , in 22 patients treated with c-Peg-V (63.9%) and in 25 patients treated with Pasireotide Lar (74.2%). These data are superimposable to those observed in previous series, as the reaching of acromegaly control is reported from 64% to 97% of patients treated with Peg-V in long term follow-up –35. ThisIn this study, we conducted a sub-analysis to evaluate the efficacy of Pasireotide Lar at 6 months of treatment, in order to rule out a possible bias due to the evaluation of efficacy of Pasireotide Lar at 12 months of treatments, that may over-estimate the control rate. We found that the efficacy of Pasireotide Lar at 6 consecutive months of treatment was around 70% and resulted superimposable to the rate of control acromegaly observed in this study in patients treated for 12 consecutive months with Pasireotide Lar (74.2%). In addition, a 6 months treatment sub-analysis confirmed that the complete resistance to first gen-SSAs was as a risk factor for the poor response to Pasireotide Lar therapy.In this study the regrowth of tumor residual occurred in one patient on treatment with m-Peg-V, one patient on treatment with c-Peg-V and one patient on treatment with Pasireotide Lar, the maintenance of first gen-SSA during treatment with Peg-V or the prescription of second generation SSA seem to be preferred in patients with tumor concern. This therapeutic approach is similar to those described in French Acrostudy, where the medical rationale for continuing SSAs rather than switching to PEG monotherapy was the tumour suprasellar extension and the reaching of tumour shrinkage during SSA . HoweverThe main limitation of this study is its retrospective design and the absence of unique treatment protocol among the different endocrine centers that enrolled patients for this study. These limitations did not allow us to provide data on the efficacy of second-line therapies, but allowed us to describe a real-life clinical experience. Moreover, according to the pathology protocol in the different centers, our study can’t provide complete data on the biology of GH secreting adenomas and on biomarkers  that may orient the choice of second-line treatments. At least, given the size and nature of our sample, it is difficult to tell whether and to what extent our observations on prescribing practices are indicative of practices in other hospitals in Italy or other countries. As for consequence, our results should be confirmed and validated in prospective studies, with larger cohort of patients, in particular with regard of the negative prognostic role of the resistance to first gen-SSA in predicting the poor outcome of Pasireotide Lar therapy, that we found in 7 patients treated for at least for 6 consecutive months with Pasireotide Lar and in 3 patients treated for 12 consecutive months.Our results confirmed that Pasireotide Lar, Pegvisomant in monotherapy or in combination with SSA are safe and effective in reaching the control of acromegaly and underline that in SSA-resistant GH secreting pituitary adenomas, c-Peg-V and Pasireotide-Lar are choose for the treatment of invasive tumors. The partial response to first gen-SSA seem to be the main determinants that guides the choice of Pasireotide Lar.Figure1SupplementaryFigure2SupplementarySupplementary Information


In addition, the clinical course of polytrauma patients with SARS-CoV-2 infection showed a high rate of pulmonary complications in the inpatient course (53%). Conclusion: The results of our study show that the changes in the CT findings of trauma patients with SARS-CoV-2 infection are a good indicator of further inpatient outcomes. Similarly, polytrauma patients with a SARS-CoV-2 infection and positive CT findings are shown to have increased risk for pulmonary complications.Background: Polytrauma patients with SARS-CoV-2 infections may be associated with an increased complication rate. The main goal of this study was to analyze the clinical course of trauma patients with COVID infection and a positive CT finding. Methods: This was a retrospective in-hospital study. Polytrauma patients diagnosed with SARS-CoV-2 infections were included in our analysis. The outcome parameters were pulmonary complication during admission, pulmonary embolism, pleural effusion, pneumonia, mortality, length of stay and readmission < 30 days. Results: 48 patients were included in the study. Trauma patients in the age-adjusted matched-pair analysis with typical changes in SARS-CoV-2 infection in CT findings showed significantly more pulmonary complications in general and significantly more cases of pneumonia (complications: 56% vs. 11%,  Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become one of the world’s most worldwide medical problems due to its global reach and the fact that more than 500 million people have already been infected. More than 5 million people have died since the start of the SARS-CoV-2 pandemic . FurtherThere are still significantly less data and guidelines regarding surgeries and emergency interventions in trauma patients with SARS-CoV-2 compared with other diseases ,4. Thus,This study is a retrospective data analysis and was approved by the local ethics committee (KEK-ZH-Nr. 2020-01037). The data for the outcome parameters were extracted from the medical records of patients in the electronic hospital information system. The data were collected over a total period of 2 years, from March 2019 to March 2021. Included were the beta, gamma, and delta variants of SARS-CoV-2. The inclusion criteria were that all patients had a positive SARS-CoV-2 test on the day of admission and trauma as their principal diagnosis at the time of admission. The test method was in every case a polymerase chain reaction (PCR) nasopharyngeal swab. All of the patients were admitted via the trauma bay or the emergency department. All patients were older than 18 years at the time of admission.Exclusion criteria were patients with a previous history of lung surgery, patients with bacterial lung diseases, patients with lung carcinomas or patients with lung carcinomas in remission, as well as when patients were referred or repatriated from other hospitals or if there was the presence of a documented refusal. Patients were divided into two groups according to the Injury Severity Score (ISS) ,12. One For the secondary study question, only the patients who were admitted via the trauma bay and consequently received a whole-body computed tomography (CT) scan were included. Again, the patients were divided into two groups: the CT (+) group and CT (−) group. In the CT (+) group, all trauma patients who were infected with SARS-CoV-2 and who had SARS-CoV-2-related changes in CT findings were included. In the CT (−) group, all trauma patients who were infected with SARS-CoV-2 but without SARS-CoV-2-related changes in CT findings were included.2 [The outcome parameters were total of pulmonary complications during admission, pulmonary embolism, pleural effusion, pneumonia, mortality during admission, length of stay, body mass index (BMI) > 25 kg/m2  and read2 , and Abb2  .It is a descriptive statistic with a retrospective data analysis. Continuous parameters were compared using Student’s A total of 48 patients with trauma as the principal diagnosis and a positive SARS-CoV-2 PCR result on admission were included in the study. There were 33 male patients (69%) and 15 female patients (31%), with a mean age of 60.3 years (SD 19.3 years). Of these patients, 15 of them (31%) were polytrauma patients with an ISS > 16. The median ISS was 10 (SD 7.0). Of all the patients, 34 of them (71%) received a whole-body CT on admission. We analyzed the pulmonary complications of polytrauma patients with SARS-CoV-2 infection on admission during their inpatient course and identified a high number of pulmonary complications (53%). There were especially high rates of pneumonia (53%) and pulmonary embolism (20%) among inpatients, as well 33% of all polytrauma patients developed a pleural effusion during their inpatient course .p = 0.001), more pneumonias (p = 0.002) and more cases of pleural effusion (p = 0.027) than in the CT (-) group  who received a whole-body CT scan we analyzed whether SARS-CoV-2 typical changes in CT findings were already present at admission . In the -) group .p = 0.141), despite changes typical of SARS-CoV-2 infection in CT findings being present. There were also no significant differences regarding length of stay (p = 0.171), mortality during admission (p = 0.210), readmission < 30 days (p = 0.952) and BMI > 25 kg/m2 (p = 0.215).There was no significant difference in pulmonary embolism (p = 0.001), an age-adjusted matched-pair analysis was performed. The age-adjusted matched-pair analysis  showed significantly more total pulmonary complications (p = 0.046) and pneumonias (p = 0.023) in the CT (+) group with typical changes in SARS-CoV-2 in CT findings. There was no longer a significant difference in pulmonary embolism (p = 0.580) and pleural effusion (p = 0.114). There were no significant differences regarding pulmonary embolism (p = 0.580), pleural effusion (p = 0.114), length of stay (p = 0.165), mortality during admission (p = 1.0), readmission < 30 days (p = 1.0) and BMI > 25 kg/m2 (p = 0.637)  .There are already many studies on guidelines for the treatment and management of severely injured patients in the inpatient setting ,16,17,18This study has shown the following main results:Polytrauma patients with SARS-CoV-2 have shown a very high complication rate . This is significantly higher compared with isolated polytrauma patients with blunt chest trauma without infection with a pneumonia rate of 12.7%  and lungp = 0.046) and pneumonias (p = 0.023) than the comparison group that had an initial CT scan of the chest that was negative for typical changes in SARS-CoV-2.It has also been shown that trauma patients with a SARS-CoV-2 infection at admission and typical changes in SARS-CoV-2 in CT findings at the primary CT scan of the chest have significantly more total pulmonary complications during admission (For the first study question, it can be concluded that it is essential to assess the risk of pulmonary embolism in polytrauma patients at a minimum from the beginning. It has already been shown in the literature that mortality is significantly higher in patients who have a SARS-CoV-2 infection in addition to a pulmonary embolism . It is aTo develop preventive measures, the pathophysiology of coronavirus thrombogenesis must first be looked at. To infect a host, the virus uses the angiotensin-converting enzyme receptors on the surface of pneumocytes in the epithelial alveolar surface . Due to In addition, care should be taken to include physical therapy in the treatment plan as early as possible in this patient population—on the one hand, to ensure early functional mobilization, which corresponds to thromboembolism prophylaxis, and on the other hand, that all these patients receive respiratory physiotherapy to prevent pneumonia.Knowing from our second study question that the trauma patients with CT findings are already at higher risk for pulmonary complications, the aim could be to avoid a second hit if possible or to keep it at least as small as possible. In concrete terms, damage control orthopedic surgery, for example, an external fixator, might be advantageous in this patient collective. However, there is a gray balance between minimizing the risk of pulmonary complications and the longer immobilization phase with delayed surgical treatment.For the daily hospital routine, this also means that, in this patient population, the required diagnostics should be performed in cases of clinical suspicion for a lung embolism as soon as possible ,32. It wThe results of this study have shown that polytrauma patients who have a concomitant SARS-CoV-2 infection and positive CT findings are highly susceptible to pulmonary complications during their inpatient course. Therefore, a major focus should be to aim for early mobilization and sufficient anticoagulation to achieve prophylaxis for pneumonia and pulmonary embolism.This was a single-center retrospective study with a small number of patients. It was difficult for a single hospital to obtain a large number of trauma patients who also tested positive for SARS-CoV-2 on the day of admission. In the future, a multicenter study would certainly be useful to obtain a larger number of patients. Additionally, the Omicron variant was not included in our study. Here, a comparison would certainly be interesting to detect potential differences.


Tight junctions in RPMI 2650 cells are opened after 30 min of incubation with SF/NAC NPs, which are demonstrated by measuring the decrease in trans-epithelial electrical resistance (TEER) values and discreteness in ZO-1 stains. The cellular uptake of SF/NAC NPs by RPMI 2650 cells is significantly greater than that for SF NPs and increased with increasing incubation time. In an in vivo imaging study (IVIS) using rats shows that the amount of NAC that is delivered to the brain by SF/NAC NPs increased by 1.40–2.60 times and NAC is retained longer in the nasal cavity than NAC solutions in a 2-h study.Using nasal routes to deliver drugs to the brain using multifunctional nanoparticles (NPs) to bypass the blood–brain barrier (BBB) might enhance the delivery efficacy. Anti-oxidative N-Acetyl-L-cysteine (NAC)-loaded silk fibroin (SF/NAC) NPs are produced, characterized and studied as a potential delivery vehicle for NAC delivered to the brain via nasal for both in vitro and in vivo studies. The NPs are not cytotoxic to RPMI 2650 cells, mucosal model cells, at a concentration of 6000 μg/mL. The anti-oxidative activities of SF/NAC NPs are demonstrated by high H Table 1In vivo intranasal delivery of FITC-NAC, which was first produced according to the procedures stated in an early study with a few modifications , to rats2/sr, ×106) for 30 min and 90 min, respectively. There was no difference between the baseline and zero for FITC-NAC that was carried by solutions because the fluorescent data had a large standard deviation. For instance, for t = 30 min and 120 min, the fluorescent intensity in the nasal cavity was 0.34 ± 0.39  and 0.75 ±1.19 , respectively. The fluorescent intensity, or the amount of FITC-NAC that was delivered to the brain of rats using SF/FITC-NAC NPs as a carrier, was about 1.40  to 2.60 times  more than that which was delivered by FITC-NAC solutions for five different sequences of measurements . The amoMore FITC-NAC was delivered to the brain (e.g., an olfactory bulb in a brain, 2O2 and a MitoSOX stain for hBMSC (SF/NAC NPs were produced by the desolvation technique, which was not cytotoxic to L929 cells at 6000 μg/mL . The hig


Data from the literature prove the procedure to be safe and effective, with promising bariatric and metabolic effects. Anyway, international societies support the creation of multicentric national and international registries to obtain more homogeneous data over the long period. We aimed to report our experience with this procedure.Biliopancreatic diversion with duodenal switch and single anastomosis duodenal-ileal bypass with sleeve gastrectomy (SADI-S) are technically demanding hypo-absorptive bariatric procedures generally indicated in super-obese patients  consenting patients were scheduled for SADI-S as primary (SADIS) or revisional procedure after sleeve gastrectomy (SADI) (respectively 87 and 34 patients). Early and late post-operative complications, operative time, post-operative stay, and follow-up data were analyzed.2 with a median age of 44 (39–51) years, the median operative time was 120 (100–155) min. Complications at 30th-day post-op were registered in 4 (3.3%) patients and late complications in 4 (3.3%) patients. At a median follow-up of 31 (14–39) months, the median percentage excess weight loss was 79.8 (55.15–91.45)%, and the median total weight loss was 57.0650 (43.3925–71.3475)%.Overall, the median preoperative BMI was 52.3 (48.75–57.05) kg/mOur data, coherently with the literature, confirm that SADI-S is a safe, effective procedure with acceptable complications rate. Larger studies with longer follow-ups are necessary to draw definitive conclusions. The duodenal switch was proposed in 1987 by DeMeester et al. , 3, creaLong-term studies have demonstrated the BPD-DS to provide significantly greater weight loss than other bariatric procedures with concurrent sustained improvement in metabolic health . HoweverLaparoscopy has become the gold standard approach for virtually all bariatric surgeries since it was first used for gastric bypass by Wittgrove et al. 1993, to reduce the invasiveness of bariatric procedures . With thTherefore, the aim of this retrospective study is to analyze our monocentric experience, that is the first in Italy, on patients undergoing SADI-S, so to support international societies in the biannual evaluation, and to encourage national and international registries to obtain a better and more homogeneous data analysis.We conducted a retrospective review of prospectively collected data into a de-identified dedicated bariatric database of patients who underwent bariatric surgery , between July 2016 and August 2021 at our tertiary referral center for bariatric surgery, Center of Excellence of the SICOb .The primary endpoint was to evaluate the safety of the procedure in terms of 30th-day complication rate. The secondary endpoint was to evaluate the safety of the procedure in terms of mid-term complications.All the patients who underwent SADI-S between July 2016 and August 2021 were identified among 2313 patients who underwent bariatric surgical procedures.Continuous variables were expressed as median (interquartile range: Q1–Q3). Dichotomic variables were expressed as number and percentage (%).For this study, follow-up was closed on the 31st January 2022.https://www.sicob.org/00_materiali/linee_guida_2016.pdf], and underwent primary (SADI-S) or revisional procedure (SADI) by laparoscopic or robotic approach. The patients were fully informed of the surgical technique, anesthesia, effects, and complications.The patients included in this study met the consensus criteria for bariatric surgery, fulfilled the national guidelines of SICOb . At discharge, the patients were advised to follow a strict dietary regimen which consists in 3 progressive phases , each one lasting at least 2–3 weeks, with proteic, vitaminic, and mineral supplementations. Proteic supplementation  is indicated because clinical practice guidelines for perioperative support of bariatric patients by the Endocrine and Obesity Societies recommend a daily intake of protein from a minimum of 60 up to 1.5 g/kg ideal body weight [https://fitforme.it/product/pacchetto-wls-maximum/#1586345186042-d5da1ce5-60ed for details of composition). FitForMe WLS Maximum® is dosed as one capsule per day. All the patients received enoxaparin (4000 UI/0.4 mL) for 4 weeks and proton-pump inhibitor (PPI)  for at least 6 months, as part of the standard post-operative protocol.A standard post-operative protocol personalized for bariatric patients was used . The sevy weight . All theThe discharge is scheduled 24 h after the surgical procedure only if the following are conditions satisfied: no clinical complications or post-operative biochemical and imaging alterations occurred; oral alimentation is tolerated; autonomy in life activities is acquired; and the discharge is accepted by the patient.Basic demographic and clinical data were collected through reviews of patient charts and electronic databases.t-test for repeated measures in parametric variables and Friedman’s test for non-parametric. The results are reported with a 95% confidence intervals.Normal distribution was assessed with the Shapiro–Wilk test. Differences in median values were analyzed with Statistical analysis was conducted with SPSS 22.0 software for Windows .This study was approved by our ethical board.With the aim of reducing the errors in the interpretation of acronyms , during the discussion we will use the term “SADI-S” to identify both primary procedures and revisional ones.During the study period, over 2313 bariatric procedures were performed . A total of 121 patients were scheduled for single anastomosis duodeno-ileal bypass with sleeve gastrectomy: 87 (71.9%) for primary procedure and 34 (29.1%) for the revisional one. The population characteristics are showed in Tables We reported 30th-day post-operative complications in 4 (3.3%) patients, and 2 of them required surgical treatment.2 who underwent laparoscopic SADIS. Right after surgery, he developed severe acute necrotizing pancreatitis of unknown origin (not biliary). Further surgery with debridement and necrosectomy was necessary (Clavien-Dindo IIIb): during this procedure, due to the severity of the abdominal situation, an anastomotic breakdown was required, and gastrostomy and nutritional jejunostomy were made. Pancreatitis was then treated with open abdomen technique and vacuum-assisted therapy with regular surgical revisions. The patient required intensive care support (Clavien-Dindo IV) for almost all the hospital stay. Gradually, we observed a clinical improvement which allowed to restore the intestinal continuity, at 4 months from first surgery, performing a one anastomosis gastric bypass (CASE 1).The first case was a 51-year-old man with a preoperative BMI 52.3 kg/m2, who underwent robotic SADIS. On POD2, the patient developed abdominal pain located on the superior umbilical trocar site and bilious vomiting after upper gastrointestinal contrast study with water soluble contrast. On physical examination, a swelling was noted on the trocar site, suspicious to be an incarcerated hernia with subileus. So urgent surgical revision (Clavien-Dindo IIIb) was mandatory, with reduction of the herniated intestinal loop and fascial closure (CASE 2).The second case was a 42-year-old woman with preoperative BMI 54.6 kg/m2 who underwent robotic SADIS. The patient developed fever on POD 2. Blood tests showed elevated acute phase reactants of inflammation. Despite negative upper gastrointestinal contrast study, the persistent fever demanded a CT scan with intravenous and oral contrast, which showed pneumonia. The patient had intravenous antimicrobial therapy (Clavien-Dindo II), following the infectious disease consultant’s indications, and her clinical condition improved rapidly (CASE 3).The third case was a 62-year-old woman with a preoperative BMI 51.4 kg/m2 who underwent robotic SADIS and was discharged on POD 2. She had uneventful course for 2 weeks after surgery, when she developed fever with elevated acute phase reactants of inflammation. An abdominal CT scan with intravenous and oral contrast was mandatory and showed a 5-cm-hematic collection near to the stomach suture line, without evidence of fistula. Consequently, the patient was readmitted and put on empirical intravenous antibiotic therapy (Clavien-Dindo II). Rapid and complete normalization of body temperature was observed (CASE 4).The fourth case was a 57-year-old woman with preoperative BMI 57.0 kg/mFour patients (3.3%) developed late complications.2 who underwent laparoscopic SADIS. He had no compliance to the dietetic regimen and to the vitaminic and oligoelement supplementation, so in the 6 following months, he developed a severe malnutrition state and a Wernicke Korsakoff syndrome. After correction of the nutritional deficits with parenteral nutrition, owing to the low compliance of the patient, we decided to turn the initial procedure into a Roux-en-Y gastric bypass (Clavien-Dindo IIIb) (CASE 5).The first patient was a 19-year-old man with preoperative BMI 58.1 kg/m2, who underwent laparoscopic SADI. One year after the initial surgery, they developed chronic diarrhea unrelated to Clostridium difficile; he recovered with antibiotic therapy per os and pre/probiotic supplementation (Clavien-Dindo II) (CASE 6). The third patient’s conditions were worse, with electrolyte imbalance and initial malnutrition due to rapid weight loss, but the patient was not willing to refer to the reference bariatric center. So, an intravenous correction of the electrolyte imbalance was necessary (Clavien-Dindo II) (CASE 7).The second and third patients were a man and a woman, 50 and 48 years old, respectively, with preoperative BMI 43.3 and 46 kg/m2, who underwent laparoscopic SADIS. At 2 years from surgery, she presented rectorrhagia, and new onset ulcerative colitis was diagnosed. Fast progression of the disease determined a malnutrition state with electrolyte imbalance that required prompt corrections. She developed also a colic perforation related to toxic megacolon, which required urgent total colectomy with end ileostomy (Clavien-Dindo IV) (CASE 8).The fourth patient was a 48-year-old woman with preoperative BMI 51.8 kg/m2, and the median daily bowel movements was 2 (2–3).At a median follow-up time of 31 (14–39) months, the median TWL  was 57.0650 (43.3925–71.3475)%; the median %EWL (percentage of excess weight loss) was 79.8 (55.15–91.45); the median BMI was 39 (25.1–36.65) kg/mIn Tables The SADI-S was described  and subsIn this study, we present the results of our SADI-S’ experiences in an Italian high volume bariatric center. We performed the first SADIS case in July 2016, and it was robotic; the first laparoscopic SADIS and the first laparoscopic SADI were performed in February 2017, and the only robotic SADI was performed in March 2021. Overall, the outcomes of 121 patients were reported. To our knowledge, this is the first monocentric Italian study on this procedure.2; the median follow-up time was 31 months; and the median %EWL was 79.8 and the median %TWL was 57.0650. These data and the other preoperative demographic characteristics are in line with the existing evidence [p = 0.233) and shorter median post-operative stay . The median operative times for robotic approach in our experience is shorter than other anecdotal experiences: Tat et al. [2, metabolic patients and revisional surgery), so we trust that it is the ideal procedure to benefit from robotic technologies.In our experience, SADI-S is a safe procedure with low post-operative complications. In the present study the starting median BMI was 52.3 kg/mevidence , 20, 22.evidence , 9, 19. evidence , we alwat et al.  reportedt et al.  reportedt et al.  experient et al. . Moreovet et al. , 29. As t et al. . The advt et al. . SADI-S We registered a 3.3% of early (30th day) post-operative complications, with 0.8% readmission rate and 1.7% reoperation rate, substantially similar to the studies of Surve et al.  and VilaThe median follow-up time is shorter than other studies , 20, andAs previously reported, the median follow-up in our study is shorter than the median follow-up to date available in the newer studies , 20: thu2 and a median %EWL of 75. Our results are comparable to those reported in the review of Spinos et al. [We performed a subgroup analysis in patients that completed 2 years of follow-up. This subgroup had a median BMI of 27.3 kg/ms et al. . Similars et al. .This is the first national monocentric retrospective study for SADI-S; however, several limitations of our analysis should be noted. First of all, this a retrospective study over a long period time. Second, the median follow-up is too short to certainly define the complication rate at middle- and long-term. Moreover, the bariatric and metabolic results over the long term have not been reported; 54.5% of patients have completed a 2-year follow-up, with only few having reached the 5-year follow-up.In conclusion, SADI-S is a safe procedure, thanks to its low rate of early complications. Our experience confirms the reproducibility of the technique, provided that it is performed in referral centers. So we can conclude that it can be one of the procedures of choice in the treatment of super-obese patient and in case of failed results after sleeve gastrectomy. However, we reckon necessary the establishment of national and international registries to obtain more homogeneous data and, most of all, to analyze the bariatric and metabolic effects over the long period in a sufficiently large number of patients, so to exclude a subpopulation bias effect.


To analyze and explore the risk factors for neurological symptoms in patients with purely hepatic Wilson's disease (WD) at diagnosis.This retrospective study was conducted at the First Affiliated Hospital of the Guangdong Pharmaceutical University on 68 patients with purely hepatic WD aged 20.6 ± 7.2 years. The physical examinations, laboratory tests, color Doppler ultrasound of the liver and spleen, and magnetic resonance imaging (MRI) of the brain were performed.The elevated alanine transaminase (ALT) and aspartate transaminase (AST) levels and 24-h urinary copper level were higher in the purely hepatic WD who developed neurological symptoms (NH-WD) group than those in the purely hepatic WD (H-WD) group. Adherence to low-copper diet, and daily oral doses of penicillamine (PCA) and zinc gluconate (ZG) were lower in the NH-WD group than those in the H-WD group. Logistic regression analysis showed that insufficient doses of PCA and ZG were associated with the development of neurological symptoms in patients with purely hepatic WD at diagnosis.The development of neurological symptoms in patients with purely hepatic WD was closely associated with insufficient doses of PCA and ZG, and the inferior efficacy of copper-chelating agents. During the course of anti-copper treatment, the patient's medical status and the efficacy of copper excretion should be closely monitored. Hepatolenticular degeneration (HLD), also known as Wilson's disease (WD), is an autosomal recessive disorder of copper metabolism. The prevalence of this disease is 1/26000 ~ 1/30000 globally, and the population carrier rate is approximately 1/90 .Symptomatic patients with WD are classified as hepatic, neurological, or mixed depending on their clinical presentation, liver imaging, liver enzymes, and magnetic resonance imaging (MRI) of the brain . The majA total of 68 patients with purely hepatic WD without neurological symptoms who were diagnosed and treated in the Department of Neurology of the First Affiliated Hospital of Guangdong Pharmaceutical University  from January 2016 to December 2021, were recruited from 65 families, including 35 men and 33 women who aged 20.6 ± 7.2 years old. All patients were followed up for more than one year. During the follow-up, neurological symptoms were assessed in 14 patients.Patients were divided into two groups, including the H-WD group  and hepatic WD with neurological symptoms at follow-up (NH-WD) group. The present study was approved by the Ethics Committee of the First Affiliated Hospital of Guangdong Pharmaceutical University. All patients signed the informed consent forms.All patients met the diagnostic criteria for WD , includiAll patients' medical records were collected, and they underwent related physical examinations and routine blood tests, in which the levels of blood alanine transaminase (ALT), blood aspartate transaminase (AST), serum copper, serum ceruloplasmin, and 24-h urinary copper were measured. All patients underwent a color Doppler ultrasound of the liver and spleen and an MRI of the brain during follow-up.Serum copper and 24-h urinary copper levels were determined by the flame method using a Hitachi Z-5000 nuclear absorption spectrophotometer .All WD patients underwent 3.0 T GE Silent MR scanning, axial T1-weighted imaging (T1WI) [repetition time (TR)/echo time (TE) = 3600 ms/100 ms], and T2-weighted imaging (T2WI) [TR/TE] = 3600 ms/100 ms], in which the layer thickness was 5.0 mm, and the layer spacing was 0.5 mm. Images were analyzed by two radiologists who had more than 10 years of experience in MRI of the brain.All patients were followed up for at least 1 year and were evaluated by routine examinations every year. The mean follow-up time was 48.9  months. Penicillamine (PCA) and zinc gluconate (ZG) were administrated as initial and maintenance therapies, respectively for all patients. PCA was administered 1–2 h before meals between 750 and 1250 mg daily, divided into 2–4 doses, and ZG was administered after meals between 210 and 420 mg daily, divided into 3 doses. The clinical, laboratory, and imaging data of all the patients were recorded and evaluated during follow-up.P < 0.05.Statistical analysis was performed using SPSS 19.0 software . Data were tested for homogeneity of variance, and normally distributed data were expressed as the mean ± standard deviation. The Howell method was used when the variance was unevenly distributed. Enumeration data were compared using the Chi-square test or Fisher's exact test, and the measurement data were compared using the independent t-test or Mann–Whitney U test. Logistic regression analysis was performed to identify risk factors for neurological symptoms. Statistical significance was set at There was no significant difference between the H-WD and the NH-WD groups in terms of gender, age, course of disease, or history of jaundice Table . BesidesP = 0.035). The color Doppler ultrasonography revealed that the number of patients with liver cirrhosis and splenomegaly was not significantly different between the two groups.There was no significant difference in the mean hemoglobin, red blood cell count, white blood cell count, and platelet count between the H-WD and the NH-WD groups. The proportion of elevated ALT and AST levels in the NH-WD group was higher than that in the H-WD group . The adherence to a low-copper diet rate was lower in the NH-WD group than that in the H-WD group (P = 0.035). The 24-h urinary copper level in the NH-WD group was higher than that in the H-WD group (P = 0.000). Moreover, the average daily oral doses of PCA and ZG in the NH-WD group were lower than those in the H-WD group .There was no significant difference in serum copper and serum ceruloplasmin levels between the two groups  patients had abnormal MRI signals of the brain. MRI of the brain in some NH-WD patients showed a remarkably high signal intensity on T1WI, high signal intensity on T2WI, and high signal intensity on diffusion-weighted imaging (DWI) .In the H-WD group, 44 (81.48%) patients had abnormal signals on MRI of the brain, and the common areas of brain damage were globus pallidus, putamen, caudate nucleus, thalamus, midbrain, and pons in 34 (62.96%), 30 (55.56%),18 (33.33%), 15 (27.78%), 6 (11.11%), and 4 (7.41%) patients, respectively. There were 32 (59.26%) patients with cerebral atrophy. There was no significant difference in the abnormal rate of MRI of the brain between the two groups  patients in the NH-WD group had abnormalities. Among 14 patients in the NH-WD group, it was revealed that 3 patients had low signal intensity on T1WI, and high signal intensity on T2WI and DWI  could have a strict low-copper diet and a regular oral intake of sufficient doses of PCA and ZG, whereas still developed neurological symptoms. It was found that patients' 24-h urinary copper levels were not high when PCA was used for anti-copper treatment. However, the 24-h urinary copper level was significantly elevated when sodium 2,3-dimercaptopropane-1-sulfonate was used for copper excretion. This suggests that oral PCA treatment for these patients is inadequate for copper excretion. Patients may receive sodium dimercaptosulphonate, dimercaptosuccinic acid, and trientine [Logistic regression analysis showed that insufficient doses of PCA and ZG influenced the neurological symptoms of patients with purely hepatic WD. The poor medication adherence may be an important reason for the inadequate dosages of PCA and ZG. In the NH-WD group, a limited number of patients (rientine  as copperientine , 26. Trerientine .The present study had some limitations. First, the retrospective design and the exclusion of patients with incomplete data might lead to bias. Second, this study was a single-center study with a relatively small sample size, which might hinder the applicability of the finding. Therefore, additional large-scale, multi-center longitudinal studies will be required to eliminate the above-mentioned limitations.In conclusion, the development of neurological symptoms in patients with purely hepatic WD was closely associated with the low doses of PCA and ZG, and the poor efficacy of individualized anti-copper therapy. In the anti-copper treatment of patients with purely hepatic WD, it is of great significance to promote patients' educational levels, monitor treatment response, and follow the principle of individualized treatment to prevent the development of neurological symptoms.


To compare the efficacy of Vonoprazan based dual treatment versus PPI based treatment for the eradication of Helicobacter pylori infection.©, m/s Getz Pharma, Pakistan) twice daily for two weeks. Confirmation of Hp eradication was done by stool Hp antigen test four weeks after completion of treatment. Nine and four patients were lost to follow-up in Group-A & B respectively. Analysis was conducted on 87 patients in Group-A and 92 patients in Group-B.A randomized controlled trial was conducted in Department of Medicine/Gastroenterology Ruth KM Pfau Civil Hospital, DMC during the period of 22 June to 21 September 2021. Sample size was calculated as 96 in each Group. All patients of age 18-75 years with Helicobacter Pylori Infection were inducted and randomly allocated to two groups. Group-A: were given Capsule Amoxicillin 1 gm; Tablet Clarithromycin 500 mg; Capsule Omeprazole 20 mg all medications were given twice daily for two weeks. Group-B were given Capsule Amoxicillin 1 gm; Tablet Vonoprazan 20 mg (Vonozonp = .042). Significantly higher frequencies of adverse events were observed in Group-A as compared to Group-B in nausea/vomiting (p = .035) and bloating (p = .045).Out of eighty-seven patients in Group-A and ninety-two patients in Group-B, 73 (83.9%) patients in Group-A and 86 (93.5%) patients in Group-B had negative H pylori result respectively after treatment (VA-dual provides an acceptable eradication rate with fewer adverse events. Helicobacter pylori (Hp) is a bacteria that causes inflammation of gastric mucosa and it is a key cause of peptic ulcer globally.Hp varies from 85-95% in developing countries and from 30-50% in developed countries.Hp should be offered eradication therapy. Multiple antibiotic regimens have been evaluated for Hp therapy. However, there are only few regimens which have consistently achieved high eradication “Initially triple therapy including a proton pump inhibitor (PPI) and two antibiotics such as amoxicillin with clarithromycin or metronidazole was considered as the best therapy for the Hp eradication”. As a result of decreased efficacy of this treatment, other different therapies have been developed.Major reason for decreased effectiveness was development of resistance to antibiotics and failure to maintain sustained increase in gastric pH of >5.7 during course of the day.1Important factors in failure of Hp therapy is local resistance to antibiotics used and if they are taken within two weeks of start of Hp eradication regimen. Clarithromycin resistance is reported at 43.9%Our objective was to compare the efficacy of Vonoprazan based dual treatment versus PPI based treatment for the eradication of Helicobacter pylori infection.Helicobacter Pylori infection: Hp infection will be labelled by any one of following investigation:Helicobacter Pylori Stool AntigenHistopathology on Giemsa StainThis Randomized control trial. was conducted in the Medical OPD and Medical Unit-1 of Dr. Ruth K.M. Pfau, Civil Hospital Karachi affiliated with Dow University of Health Sciences from 22 June 2021 till 21 September 2021was Non-Probability ConsecutiveGroup sample sizes of 96 in Group-A and 96 in Group-B achieve 95.246% power to detect a difference between the group proportions of 0.3. The proportion in Group-A is assumed to be 0.5 under the null hypothesis and 0.8 under the alternative hypothesis. The proportion in Group-B is 0.5. The test statistic used is the two-sided Z-Test with unpolled variance. The significance level of the test is 0.05”. Calculation was done using PASS 2019 Power Analysis and Sample Size Software . The Sampling Technique: All patients between age of 18-75 years with Hp infection as described above.History of gastric surgery such as partial gastrectomy.History of allergy to drugs used in the study.Intake of antibiotics, PPIs, corticosteroids within the last two weeks.Pregnant or lactating females.Alcohol or drug addiction.Severe neurological or psychiatric disorder.All patients presenting to the OPD or Medical Unit -I of Dr. Ruth KM Pfau, Civil Hospital Karachi and fulfilling the inclusion criteria were included after taking informed consent.The participants were randomly allocated to two groups with help of random number table and were treated for two weeks.Capsule Amoxicillin 1 gm twice dailyTablet Clarithromycin 500 mg twice a dayCapsule Omeprazole 20 mg twice a dayCapsule Amoxicillin 1 gm twice daily©, m/s Getz Pharma, Pakistan).Tablet Vonoprazan 20 mg twice a day  of age was compared between groups using χTwo hundred thirty-three patients were assessed for eligibility out of these a total of 41 patients were excluded. The breakup of these was, not fulfilling selection criteria (n=22), declined to give consent (n=16) and history of using PPI in preceding two weeks (n=3). Randomization was continued till sample size of 96 in each group was achieved. Nine patients were lost to follow-up in Group-A, while four patients were lost to follow-up in Group-B. Analysis was conducted on 87 patients in Group-A and 92 patients in Group-B. Details of enrollment & randomization are given in 2  = .09; p = .754. Mean age ±SD in Group-A was 40.2 ±9.8 years and in Group-B was 41.4 ±10.6 years respectively, statistically there was no significant difference in age between two groups t (177) = -0.81, p = .422.Total one hundred seventy-nine patients were analyzed which included 109 males and 70 females. In Group-A there were 54 (62.1%) males and 33 (37.9%) females and in Group-B there were 55 (59.8%) males and 37 (40.2%) females. Both groups were comparable in frequency of gender c2  = 4.126; p = .042. Vonoprazan based dual therapy more likely to eradicate H. pylori than convention PPI based triple therapy.Out of eighty-seven patients in Group-A and ninety-two patients in Group-B, 73 (83.9%) patients in Group-A and 86 (93.5%) patients in Group-B had negative H pylori result respectively after treatment, cFew adverse effects were reported, but none so severe to stop treatment. Significantly higher frequencies of adverse events were observed in Group-A as compared to Group-B in nausea/vomiting and bloating, details are given in To the best of our knowledge, this is the first randomized control study to show the efficacy and safety of 14 days Vonoprazan amoxicillin dual therapy. Results of our study demonstrated higher H. Pylori eradication rate of 93.5% by Vonoprazan based dual therapy as compared to PPI based conventional triple therapy, which achieved 83.9% eradication rate (p<0.05). In majority of trials a 7-day dual or triple therapy of vonoprazan was given along with either amoxicillin alone or amoxicillin plus clarithromycin to reveal the eradication rate upto 85% to 87%.et al., in their meta-analysis on efficacy and safety of VPZ-based versus proton pump inhibitor-based triple therapy for helicobacter pylori eradication, reported significantly less incidence of adverse effects and well tolerability in VPZ-based triple therapy than that in PPI-based triple therapy .9Another important finding in our study was fewer adverse effects and easy tolerability in vonoprazan based dual therapy compared to conventional triple therapy. Only few patients in VPZ group developed nausea/ vomiting, bloating and diarrhea, but none was severe enough to stop regimen. Similarly Qiu-Ju Lyu +,K+-ATPase is the preferred target for acid suppression. Until recently, the only drug that effectively inhibited this ATPase was the proton pump inhibitor (PPI). PPI is a pro drug, that require acid for its activation. Once acid-activated, PPI bind to ATPase (proton pump) to inhibit acid secretion. But because of short half-life of PPI and continued de novo production of the H+, K+-ATPase, night-time acid secretion is not fully inhibited. Whereas vonoprazan, which is a potassium-competitive acid blocker (PCAB) is an active drug that does not require acid for its activation, has seven hours half-life, bind reversibly with both active H+, K+-ATPase pumps and Inactive pumps with slow dissociation rate. PPI require meal for its activation while Vonoprazan has no meal dependency. Onset of action of Vonoprazan is within 24 hours, pH >4 sustained over most of the time, including overnight in majority of patients, without tolerance to the drug.Although Vonoprazan does not have anti-microbial properties, this significant superiority of Vonoprazan over PPI for H-Pylori eradication is suggested mainly from more sustained maintenance of gastric PH as VPZ 40 mg daily can maintain pH > 4.0 for > 90% of the day.et al. in his study brought up the fact that vonoprazan and amoxicillin dual therapy achieved a cure rate of 80%, when given in clarithromycin resistant patients.Antimicrobial stewardship and recognizing optimal treatment and duration of therapy, without unjustified use of antibiotics is imperative in current era of antimicrobial resistance. DY Graham One of the main strengths of our study is its study design of being randomized control that provides higher level of statistical reliability. A meta-analysis, where 16 studies were selected for quantitative review, had limitation that 15 of 16 studies in their meta-analysis were retrospective and used historical controls with a time-frame shift between the two groups,In the era of growing antimicrobial resistance, VA-dual therapy is a potential new first- line Hp therapy for cases because it provides an acceptable eradication rate and high safety and will have a potentially less negative impact on future antimicrobial resistance.BFZ, FSA, NB: Substantial contributions to conception and design, or acquisition of data, or analysis and interpretation of data. TR, SMH, AS: Drafting the article or revising it critically for important intellectual content BFZ: Final approval of the version to be published. NB: Statistical Analysis. Agreement to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. (All Authors).


APA parameters showed moderate correlation with the MDS-UPDRS Rigidity, Characterizing-FOG questionnaire and FAB-2 planning. The single IMU-based reconstruction algorithm was effective in measuring APAs timings in PD. The current work sets the stage for future developments of tele-rehabilitation and home-based exercises.Out-of-the-lab instrumented gait testing focuses on steady-state gait and usually does not include gait initiation (GI) measures. GI involves Anticipatory Postural Adjustments (APAs), which propel the center of mass (COM) forward and laterally before the first step. These movements are impaired in persons with Parkinson’s disease (PD), contributing to their pathological gait. The use of a simple GI testing system, outside the lab, would allow improving gait rehabilitation of PD patients. Here, we evaluated the metrological quality of using a single inertial measurement unit for APA detection as compared with the use of a gold-standard system, i.e., the force platforms. Twenty-five PD and eight elderly subjects (ELD) were asked to initiate gait in response to auditory stimuli while wearing an IMU on the trunk. Temporal parameters  extracted from the accelerometric data and force platforms were significantly correlated (mean(SD), r: 0.99(0.01), slope: 0.97(0.02)) showing a good level of agreement (LOA [s]: 0.04(0.01), CV [%]: 2.9(1.7)). PD showed longer APA-Duration compared to ELD ([s] 0.81(0.17) vs. 0.59(0.09)  Gait initiation (GI) is a complex transient task performed between the quiet standing posture and steady state walking, requiring a shift from a static stable state to a, relatively less stable, dynamic state of movement . For thiAPAs have been extensively studied in persons with Parkinson’s disease (PD), because of their balance deficits, gait impairments and frequent falls . PD is cTo address this gap, Gazit et al.  developeIn consideration of the limitation of using the multiple wearable sensors approaches reported above, the twofold aim of this work was to evaluate: (1) if the acceleration-based algorithm described in , that neTwenty-five subjects with mild-to-moderate idiopathic PD (thirteen without and twelve with previous experiences of FOG)  and eighConsidering the main aim of the present study , the sample size calculation was estimated using previous published data related to the Bland–Altman mean difference (GRF vs. IMUs) of the APA duration parameters. Bonora et al.  found a Disease severity was evaluated with the section III of the MDS–Unified Parkinson Disease Rating Scale . The patThe clinical assessment included the administration of the following clinical scales: the short physical performance battery , the FoParticipants stood on the first force platform with their feet in parallel at hip-width and were asked to initiate a gait in response to auditory stimuli provided through a headphones set and to stop as soon as possible with both feet on the second force platform. The protocol included five trials and the auditory stimulus was a neutral voice saying ‘go’. No indication was given on the starting foot. https://www.cometasystems.com/), equipped with a MEMS sensor . The device has a low weight (5.3 g) and small size (32 mm × 24 mm × 7 mm), is waterproof, and is equipped with a wireless interface for real-time data streaming, as well as 1 GB of on-board memory which allows to store locally up to 6 h of continuous measurements. For the study purposes, only data of the triaxial accelerometer were analyzed at a sampling rate of 140 Hz with a 16-bits resolution. The sensor was fixed with an elastic band on the trunk at L5 level. The acceleration signal was processed using a 4th-order, band pass Butterworth filter between 0.2 and 4.5 Hz.The wearable measurement system used consists of a single IMU with a 16-bits resolution, analog-to-digital converter and a full scale of ±16 g  displacement were measured with the force platforms, considered as gold standard, at 1000 Hz . Retroreflective markers were placed on malleolar anatomical landmarks . COP displacements recorded from the force platforms were filtered with a fourth-order, zero-lag, low-pass Butterworth filter with a cut-off frequency of 10 Hz.A custom-made Software, developed in Visual Studio dot net  environment, recorded the accelerometric data and controlled the trial onset and offset, the auditory stimulus presentation and the synchronization with the optoelectronic system including the GRFs. For each stimulus sent to the headphones, a synchronous trigger signal was sent to SMART Capture software .APAs timings quantification were calculated from three automatically-detected time points: (1) APA onset, 2) toe-off, and 3) heel-strike both from the CoP data : 73.9 (6.2), body mass [kg]: 68.4 (10.8), body height [cm]: 169.0 (6.0)); ELD 5 Males, mean (SD), age [yrs]: 68.9 (7.5), body mass [kg]: 70.6 (13.0), body height [cm]: 169.1 (6.8), A good agreement of the IMUs compared to the reference system was observed for the APA timing measures through both the Bland–Altman method and linear correlation B–F. Therp = 0.50), toe-off  and heel-strike  instants. Moreover, for the MAEs values of the parameters APA and Swing durations, no significant differences were found .No significant differences were found between PD and ELD subjects considering MAEs for APA onset  uses to maintain and restore balance when the equilibrium state is perturbed or changed . Human vConcerning the sensitivity, GI metrics extracted from a single IMU were able to discriminate between PD and elderly on the basis of APAs alterations. More precisely, persons with PD showed prolonged APAs compared to those of elders during GI . Thus, the ability of the algorithm to detect differences in more homogeneous groups should be further investigated.The acceleration -based algorithm tested in this study is a promising tool for assessing GI of persons with Parkinson’s Disease. Results of the validity procedure conducted in this study demonstrated a strong agreement between a single IMU-based measurement system and the platforms acquisition system to estimate the APAs timings. In addition, the GI metrics were associated with clinical features of persons with PD, being able to discriminate between persons with PD who experienced freezing of gait and those that did not. APA impairment is a common feature among subjects with different neurological diseases and, therefore, future studies to validate the method on subjects affected by other neurological disorders, such as Multiple Sclerosis, are encouraged. In fact, a recent study highlighted that in the early stage of multiple sclerosis, although the APA of GI are strongly affected, there are no significant alterations of the executive phase , suggest


Intestinal fibrosis, one of the complications of inflammatory bowel disease (IBD), is associated with fistula and intestinal stricture formation. There are currently no treatments for fibrosis. Mesenchymal stem cell-derived exosomes have been proven to exert inhibitory and reversal effects in IBD and other organ fibrosis. In this study, we explored the role of human umbilical cord mesenchymal stem cell-derived exosomes (hucMSC-Ex) in IBD-related fibrosis and its associated mechanism to provide new ideas for the prevention and treatment of IBD-related intestinal fibrosis.  We established a DSS-induced mouse IBD-related intestinal fibrosis model and observed the effect of hucMSC-Ex on the mouse model. We also used the TGF-induced human intestinal fibroblast CCD-18Co to observe the role of hucMSC-Ex in the proliferation, migration, and activation of intestinal fibroblasts. Having observed that the extracellular-signal-regulated kinase (ERK) pathway in intestinal fibrosis can be inhibited by hucMSC-Ex, we treated intestinal fibroblasts with an ERK inhibitor to emphasize the potential target of ERK phosphorylation in the treatment of IBD-associated intestinal fibrosis. β-induced proliferation, migration, and activation of human intestinal fibroblasts, and ERK phosphorylation played a key role in IBD-associated fibrosis. The inhibition of ERK decreased the expression of fibrosis-related indicators such as α-SMA, fibronectin, and collagen I.  In the animal model of IBD-related fibrosis, hucMSC-Ex alleviated inflammation-related fibrosis as evident in the thinning of the mice's intestinal wall and decreased expression of related molecules. Moreover, hucMSC-Ex inhibited TGF- hucMSC-Ex alleviates DSS-induced IBD-related intestinal fibrosis by inhibiting profibrotic molecules and intestinal fibroblast proliferation and migration by decreasing ERK phosphorylation. Inflammation bowel disease (IBD) is a severe intestinal inflammation, including ulcerative colitis (UC) and Crohn's disease (CD). Intestinal fibrosis is a complication in IBD, especially in CD, which triggers the occurrence of stricture in the locations involved. Apart from stricture in the intestinal lumen, IBD-related fibrosis can induce anal fistula, an important complication in CD, when the inflammation appears in the rectum .β (TGF-β) is a key molecule in the progression of intestinal fibrosis. TGF-β functions in fibrosis by influencing downstream signaling pathways such as the TGF-β/smad pathway and the wnt/β-catenin pathway [The mechanism of intestinal fibrosis is not completely confirmed. In a recent study, it is widely accepted that transforming growth factor- pathway . Several pathway .In addition to not fully understanding the mechanism involved in intestinal fibrosis, there is also no ideal therapeutic method. Although some of the existing anti-inflammation drugs have been shown to prevent fibrosis to a certain extent, those therapies are not always useful as antifibrotic. Mesenchymal stem cells (MSCs) have the ability to self-renew and multidifferentiate . There aα and IL-1β and increasing the level of IL-10 [Exosomes are small vesicles secreted by cells and are considered to be a type of extracellular vesicle (EV). They have a diameter between 30 and 150 nm and a density between 1.13 and 1.19 g/ml . For exaof IL-10 . Moreoveof IL-10 . RegardlTherefore, in this study, we explored the role and mechanism of hucMSC-Ex in mitigating IBD-related intestinal fibrosis and provide new insights into intestinal fibrosis treatment approaches.The study was approved by the Ethical Committee of Jiangsu University (2012258).2 environment, a small amount of α-MEM (Invitrogen) containing 15% FBS  and 5% penicillin-streptomycin (Invitrogen) was used to cover segments in dishes, and the solution was replaced every 3 days. When observation under the microscope revealed long spindle cells near the small pieces of tissues and fused to 80%, it indicates that hucMSCs could be digested.Fresh umbilical cords were obtained from the Zhenjiang Fourth People's Hospital. The umbilical cords were washed in PBS for 30 minutes and were then cut into pieces and placed in clean 3.5 cm small dishes to adhere to the wall. After incubation at 37°C in a 5% COα-MEM with 10% fetal bovine serum  and 1% penicillin-streptomycin (Invitrogen). When the hucMSCs in the Petri dish grew to about 70% confluence, the nutrient solution was replaced by a new α-MEM (Invitrogen) nutrient solution, which is made up of 10% exosome-free FBS and 5% penicillin-streptomycin (Invitrogen). After culturing in the incubator for 48 hours, the supernatant was collected and the above process was repeated until the P7 passage. Human intestinal fibroblasts CCD-18Co were purchased from Beiner Biotechnology and cultured in RPMI 1640 medium (Invitrogen), which constitutes 10% FBS  and 5% penicillin-streptomycin (Invitrogen).hucMSCs were isolated and cultured in μm filter and distributed into EP tubes for storage in a -80°C refrigerator.hucMSC-Ex was extracted by ultracentrifugation. The supernatant of hucMSC culture was centrifuged at 4°C, 2000g for 30 minutes. The supernatant of the previous step was centrifuged at 4°C, 10000g for 30 minutes. Next, the obtained supernatant was concentrated to 100 ml using 100KD ultrafiltration tubes. The concentrated liquid was further centrifuged at 4°C, 100,000g for 70 minutes. The resultant precipitate was saved, and an appropriate amount of PBS was added to dissolve it. The dissolved solution was centrifuged at 4°C, 100,000g for 70 minutes, the precipitate was collected, and an appropriate amount of PBS was added to dissolve the precipitate. Finally, the resultant solution was filtered with a 0.22 The standard solution was used to generate a standard curve, and diluted hucMSC-Ex was added to the sample chamber and read the data. Use the software for image capture and analysis.μl hucMSC-Ex solution was dripped onto a 2 mm copper mesh. The copper mesh was allowed to stand, and excess liquid was wiped off the edge. hucMSC-Ex was counterstained with 3% phosphotungstic acid, and pictures were taken with the transmission electron microscope after drying.About 20 2O), while the DSS group was given 2% DSS solution throughout the first week and two weeks later and given food for three cycles and ddH2O when 2% DSS solution is not being administered. The hucMSC-Ex group received 100 μl (1 × 109 particles) hucMSC-Ex by tail vein injection on the 3rd, 10th, 17th, 24th, 31st, 38th, 45th, 52nd, and 59th days. All mice were weighed every 3 days, and fecal appearance and consistency were observed. On the 63rd day, the mice were sacrificed.The study used 6-8-week-old female C57BL/6 mice purchased from the Animal Experiment Center of Jiangsu University. The mice were weighed and randomly divided into three groups: NC group, DSS group, and hucMSC-Ex group Sup. . The NC The disease activity index (DAI) was calculated based on the criteria shown in Staining kits (Solarbio) were used for hematoxylin and eosin (H&E), Masson, and Sirius red staining of paraffin sections of mouse intestinal tissues.β-actin , anti-α-SMA , anti-collagen I , anti-collagen III , anti-fibronectin , and anti-t-ERK  overnight. After washing with 1x TBS/T buffer, the membranes were incubated with the secondary antibody  for 30 min at 37°C. Pictures were taken with a chemical gel imaging system (GE).A moderate volume of RIPA solution was added to cells and mixed thoroughly. The solution was shaken on a vortex shaker for 1 minute and allowed to stand on ice for 5 minutes. The process was repeated 5 times in total. The solution was centrifuged at 4°C, 12,000g for 15 minutes to obtain the supernatant, which is the protein solution. SDS-PAGE electrophoresis was used to separate target proteins and transfer them to a PVDF membrane. The PVDF membranes were soaked in 5% skim milk to block nonspecific proteins. The membranes were incubated with corresponding primary antibodies: anti-CD9 , anti-CD81 , anti-HSP70 , anti-calnexin , anti-RNA was extracted by TRIzol (Vazyme) and dissolved in DEPC water. cDNA was obtained by using the Vazyme kit on the PCR machine . Obtained cDNA was used to perform QRT-PCR using the Vazyme kit on Step One Plus Real-Time PCR System (ABI). The reaction process was 95°C, 5 minutes; 95°C, 30 seconds; 58~60°C (flexible setting according to different primers), 30 seconds; and 72°C, 30 seconds, for a total of 40 cycles. The specific system reagents and volumes are shown in Tables Mouse colon tissues were fixed in 4% paraformaldehyde. The tissues were dehydrated, cleared, and embedded in paraffin, followed by microtome sectioning at the First People's Hospital of Zhenjiang City. The slices were kept at 56°C for 4 hours to prevent the tissue from falling off in subsequent procedures. The tissue slices were deparaffinized, and the sections were placed in a hydrogen oxide-methanol solution (1 : 9 mixed with 30% hydrogen peroxide and methanol) and kept at room temperature for 30 minutes. Slices were washed 3 times with histochemical PBS, immersed in citrate solution, and steamed in an electric oven for 30 minutes. After cooling, the histochemical PBS was washed 3 times, followed by blocking nonspecific proteins in sections with BSA for at least 30 minutes. The protein antibody (Proteintech) was diluted to 1 : 200 with 5% BSA and used to cover the tissue at 4°C overnight. It was then washed 3 times with PBS, incubated with the secondary antibody  and SABC , and finally observed under the microscope after adding DAB (Bio-Rad), looking for golden yellow positive areas. The hematoxylin staining solution was used to counterstain the slide, and pictures were taken under the microscope.β-induced cell models  nutrient solution was added to the well. Three groups of cells in the six-well plate were digested and resuspended in FBS-free RPMI 1640 nutrient solution. A certain amount of resuspension was added to different Transwell wells, and the 24-well plate was cultured at 37°C and 5% CO2 for 18 hours. Cells were stained with 0.5% crystal violet, and images were obtained under the microscope.The TGF-els Sup.  were conβ  and anti-fibronectin  overnight. A fluorescent secondary antibody (SAB) was added to the slide and incubate for 1 hour at 37°C in the dark and then washed with PBS 3 times. Nuclei were stained with Hoechst 33342, the slides were fixed with an antifluorescence quencher, and images were taken under the laser scanning confocal microscope.Cell slides were placed in a 24-well plate, and cells were added to construct a cell model induced by TGF-β Sup. . After tP values of < 0.05 were considered statistically significant.Data for all results were shown as mean ± standard error. Analysis of variance was performed by using the GraphPad Prism software, and 10/ml.The extracted hucMSC-Ex were examined with nanoparticle tracking analysis (NTA), which revealed particle diameter between 50 and 150 nm . Transmiμl (1 × 109 particles) of hucMSC-Ex via the tail vein on the third day of every week, starting from the first week. The results showed variations in the weight of the different groups. With time, the weight of mice in the NC group gradually increased while that of the DSS group showed a downward trend with the administration cycle. However, body weight loss was effectively suppressed after the hucMSC-Ex injection  group, a DSS group, and a hucMSC-Ex group. The hucMSC-Ex group received 100 njection . At the njection . On day njection . To assenjection .β, and α-SMA was down-regulated in the hucMSC-Ex group . However, the expression of COL3A1 genes increased in the TGF-β group, and the changes were not obvious or even continued to increase after hucMSC-Ex treatment  and ERK phosphorylation. Moreover, treatment of TGF-β-induced fibroblasts with ERK phosphorylation inhibitors showed that the expression of α-SMA, collagen I, and fibronectin decreased. All the above results indicated that hucMSC-Ex could inhibit the ERK phosphorylation level in fibroblasts and thus inhibit the development of fibrosis.ERK is the final component of the Raf-MEK-ERK signaling module, which acts downstream of receptor tyrosine kinases and controls multiple cellular processes, including cell proliferation and differentiation. Activated bi-phosphorylated ERKs transmit pathway activation to cells through the phosphorylation of multiple substrates. The ERK pathway plays a regulatory role in a variety of cellular activities. Abnormalities in the ERK pathway could cause neurodevelopmental disorders. On one hand, ERK activation causes the proliferation of cancer cells. ERK has a proapoptotic effect, and enhanced ERK1/2 signaling can lead to tumor cell death . In addire aging . TherefoThis study demonstrates that exosomes can affect the occurrence of IBD-associated intestinal fibrosis by regulating ERK phosphorylation in human intestinal fibroblasts. However, our study has shortcomings, including the lack of a detailed mechanism of ERK phosphorylation in fibrosis inhibition and the use of only intestinal fibroblasts in the ERK inhibitor PD98059 experiment, which makes the evidence weak. In addition, there is a lack of data on clinical samples. There is still some gap between the simulated disease occurrence process and the real disease occurrence; thus, clinical studies are needed, which could lead to the safe use of exosomes in humans for IBD-related intestinal fibrosis treatment. Many cells play a role in the development of fibrosis. Studies have proved that hucMSC-Ex can regulate the proliferation and activation of intestinal fibroblasts. Whether other intestinal cells can also be regulated by hucMSC-Ex can also be further investigated. Nonetheless, our findings are significant and inspire further exploration in this area. We expect these findings to be linked to previous results in other organ fibrosis and IBD and provide new insights for inhibiting and reversing fibrosis.β-induced intestinal fibroblast proliferation, migration, and activation. The hucMSC-Ex relieve-effect is associated with its inhibition of ERK phosphorylation in intestinal fibroblasts, leading to reduced expression of fibrotic molecules to alleviate intestinal fibrosis.hucMSC-Ex alleviates DSS-induced IBD-related intestinal fibrosis in mice by inhibiting TGF-